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Functional compartmentation of DNA precursors in human leukaemoblastoid cell lines
Author(s) -
Taheri M. R.,
Wickremasinghe R. G.,
Hoffbrand A. V.
Publication year - 1982
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.1982.tb03909.x
Subject(s) - thymine , nucleotide , cytosine , thymidine , dna , intracellular , biochemistry , guanine , microbiology and biotechnology , cell culture , hl60 , chemistry , biology , cell , genetics , gene
S ummary . Human leukaemoblastoid cell lines were incubated with [ 3 H]t hymidine and used to study the utilization for DNA synthesis and degradation of thymine nucleotides labelled via the salvage pathway. Three types of experiments were performed. (1) Cells were continuously labelled with exogenous [3H]thymidine at 3 7°C to compare the incorporation of label into DNA with the specific radioactivities of thymine nucleotides separated by chromatography on polyethyleneimine‐cellulose. These studies demonstrated whether one or more functional pool of thymine nucleotides was present. (2) Intracellular thymine nucleotide pools were labelled with [ 3 H]thymidine at 13°C, and the cells were subsequently incubated in unlabelled medium at 37°C in order to quantify the percentage of thymine nucleotides incorporated into DNA. The results showed that 18%, 75%, 75% and 91% of intracellular thymine nucleotides were incorporated into DNA during the chase, in Raji (B lymphoid), Nalm‐6 (pre‐B cell), HL60 (myeloid) and KM3 (c‐ALL) cells respectively, whereas Molt4 (Thy‐ALL) cells incorporated 100%. (3) In order to quantify the activity of the degradative pathway for thymine nucleotides in these cell lines, cells were labelled as described in (2) in the presence of cytosine arabinoside (an inhibitor of DNA synthesis). 100%, 90%, 65%, 38% and 20% of labelled thymine nucleotides were degraded in Raji, HL60, Nalm‐6, KM3 and Molt‐4 respectively. These results confirm our previous conclusion (Taheri et al, 1981a, b) that thymine nucleotides are functionally compartmentalized in human cells. If these results with cell lines can be extrapolated to apply to de novo leukaemia cases, they suggest that the degree of compartmentation differs widely between different leukaemia cell types. Thy‐ALL cells utilized all their labelled thymine nucleotides for DNA synthesis, whereas B‐ALL, cells utilized only 18% of their labelled thymine nucleotide nucleotides. Thy‐ALL cells have the smallest degradative thymine pool and degradative capacity whereas B‐ALL cells possess the largest degradative pool and capacity. AML, pre‐B‐ALL, c‐ALL cell lines show intermediate results. If these results reflect those of the normal bone marrow cells from which each of the leukaemias arise, they suggest that with progressive B cell development in the bone marrow, degradative ability for DNA precursors increases and the efficiency of utilization of DNA precursors for DNA synthesis decreases.